The contents of all patents and publications cited in this specification are hereby incorporated by reference in their entirety.
Hydrogels are highly hydrated, macromolecular networks, dispersed in water or other biological fluids. Hydrogels that exhibit the specific property of increased viscosity with increased temperatures are known as thermoreversible, thermosensitive (or thermosetting) hydrogels. It is known that thermosensitive hydrogels may be prepared from polymers of natural origin such as chitosan, which is a commercially available, inexpensive polymer obtained by partial to substantial alkaline N-deacetylation of chitin, a linear polysaccharide, made of N-acetylglucosamine units, linked via β-1,4-glycosidic bonds. The deacetylation process is generally performed using hot, concentrated, hydroxide solutions, usually sodium hydroxide.
Chitosan is biocompatible, non-toxic, and non-immunogenic, allowing its use in the medical, pharmaceutical, cosmetic and tissue construction fields. For example, topical ocular applications and intraocular injections or transplantation in the vicinity of the retina have been used. Moreover, chitosan is metabolized-cleaved by certain specific enzymes, e.g., lysozyme, and can therefore be considered as biodegradable. In addition, it has been reported that chitosan acts as a penetration enhancer by opening epithelial tight junctions. Chitosan also promotes wound healing and exhibits antibacterial, antifungal and antitumor properties.
The complexity of biological structures such as natural tissue has resulted in researchers exploring the use of biomaterials and medical devices that are introduced on the skin or into the body of a subject as a liquid and that turn solid or solid-like through simple application or injection. For example, chitosan hydrogels have been shown to be useful for cartilage regeneration and prevention of knee pain associated with acute and chronic cartilage defects. Chitosan-based gels have also been shown to turn into and serve as scaffolds for the encapsulation of invertebral disc (IVD) cells by entrapping large quantities of newly synthesized anionic proteoglycan. Chitosan is known to form thermoreversible gels in the presence of several multivalent anions, such as phosphate derivatives. Temperature-controlled pH-dependent formation of ionic polysaccharide gels, such as chitosan/organo-phosphate aqueous systems, has been described, for example, in PCT International Publication No. WO 99/07416 and U.S. Pat. No. 6,344,488. However, hydrogels made from ionic polysaccharides such as chitosan are weak and usually form only after a relatively long waiting time, after mixing polymer and salt solution. This is mainly due to the fact that it is difficult to obtain homogenous, fully-hydrated chitosan solutions with a high concentration of chitosan, especially high molecular weight chitosan, due to its poor solubility. Further, several medical applications require provision of not only a simple sol-gel transition, but a solid structure with desired macroporosity and mechanical properties. Moreover, temperature is a non-specific stimulus and can be triggered by elements outside the human body such as hot weather or, for oral applications, simply drinking a hot beverage. Thus, there is a need for stimuli-responsive implants and patches that can reach desired mechanical and/or permeability properties only when triggered by specific physiological stimuli.
U.S. Pat. No. 9,034,348 discloses injectable chitosan mixtures forming hydrogels. There are described chitosan compositions which form a hydrogel at near physiological pH and 37° C., comprising at least one type of chitosan having a degree of acetylation in the range of from about 30% to about 60%, and at least one type of chitosan having a degree of deacetylation of at least about 70%. Further disclosed is a chitosan composition which forms a hydrogel at near physiological pH and 37° C. that includes at least one type of chitosan having a degree of deacetylation of at least about 70% and a molecular weight of from 10-4000 kDa, and at least one type of a chitosan having a molecular weight of from 200-20000 Da. Also disclosed are methods of preparation and uses of the chitosan compositions.
U.S. Patent Application Publication No. 2010/0028434 discloses temperature controlled and pH-dependent self-gelling biopolymeric aqueous solutions. There are described biopolymeric liquid aqueous compositions for producing self-gelling systems and gels, which comprises an acidic water-based medium, 0.1 to 10% by weight of a pH-gelling acid-soluble biopolymer, and 0.1 to 10% by weight of a water-soluble molecule having a basic character and a pKa between 6.0 and 8.4, or a water-soluble residue or sequence of the molecule having a basic character and a pKa between 6.0 and 8.4. The liquid compositions have a final pH ranging from 5.8 and 7.4, and form a stable solid and homogeneous gel within a temperature range from 10 to 70° C. Methods for preparing the compositions and uses thereof are also described.
U.S. Patent Application Publication No. 2010/0285113 discloses inverse thermal gelling composite hydrogels having enhanced stability. There are described composite hydrogels comprising a blend of an aqueous solution of an anionic polysaccharide or a derivative thereof, such as hyaluronan (also commonly referred to as hyaluronic acid) or a derivative thereof and an aqueous solution of methylcellulose or another water soluble cellulose derivative thereof, having dispersed polymeric particles, such as polymeric hydrophobic particles therein selected from microparticles and nanoparticles, and wherein the stability of the hydrogel is enhanced relative to the stability of the hydrogel alone. The polymeric particles may contain at least one therapeutic agent, in which case each therapeutic agent exhibits a linear sustained release rate that can be tuned or altered by selecting the appropriate polymer formulation of the microparticles and/or nanoparticles. The composite may be injectable, and in the absence of a therapeutic agent may be used as a bulking agent for reconstructive and cosmetic surgery or may act as a platform for subsequent delivery of therapeutic agents.
Insofar as veterinary health issues are concerned, mastitis is an inflammation of the mammary gland that is typically caused by bacteria which in most cases enter the gland via the teat orifice. During the non-lactating period or “dry period” in the gland, deposits of keratin in the teat orifice and the streak canal form a primary defense mechanism. A keratin plug that forms in the teat of the animal forms a protective barrier, and the immune-rich tissues of the Furstenburg's Rosette in the teat, as well as the natural protective factors of the dry-cow secretions, contain high levels of naturally occurring anti-bacterial substances (cationic proteins) which inhibit the passage of bacteria from the teat orifice to the teat cistern (papillary sinus) and gland cistern. However, this keratin plug and these natural immune defense mechanisms can be overcome by bacterial invasion as the animal enters into the dry period at the end of lactation, during the dry period of the animal, and/or during calving. As a result, bacteria invade the gland and cause mastitis during the dry period or, more particularly, immediately following calving.
The major pathogens causing mastitis are Staphylococcal species such as, for example, Streptococcus agalactiae, Staphylococcus aureus and the like, Corynebacterium bovis, Mycoplasma, coliforms such as, for example, Esherichia coli, Klebsiella spp., Enterobacter spp., and Citrobacter spp., environmental Streptococcal species such as, for example, Strep. dysgalactiae, Strep. uberis, and Enterococcus spp., Pseudomonas spp., etc. Although mastitis is mainly caused by bacteria, the inflammation can also be produced as a result of viral infection (e.g., bovine herpesvirus II and IV, a paravaccinia virus such as Pseudo Cowpox, and the like) or infection with atypical pathogens like mycotic (e.g., Candida spp. and Aspergillus spp.) or algal microbes (e.g., Prototheca spp.) with or without development of a secondary bacterial infection.
Mastitis due to the presence of pathogens can become a highly contagious condition within the confines of a dairy farm that results in huge production losses for the dairy industry. Reduction of drinkable milk then occurs from the harmful pathogens' effects or various treatments that render the milk not fit for human consumption. While severe cases can end in death, unhindered outbreaks can also cause permanent damage to the animals' udders. As a major endemic disease of dairy animals, mastitis puts the animal welfare at risk and often entails rather costly veterinary care. The value of protecting the early lactation period from existing and new infections perpetuated from the dry period remains highly valuable to the industry. It is clear that the treatment and control of mastitis is an important goal to maintain the animal's health and to lower the high costs of milk production in the dairy industry.
To that end, products have been developed in an attempt to seal an animal's teat to prevent mastitis and other conditions, for example, barrier teat dips to seal the external surface and streak canal of the teat during periods of milking and internal teat sealants to block or to seal the teat canal or to plug the teat cistern during the dry period, especially for heifers and cows that have experienced one or more pregnancies previously.
Along with these products, several methods to reduce the incidence of mastitis are described in the art, for example, a method comprising sequentially delivering from a single delivery device an antimicrobial formulation and a seal formulation into the teat canal of a non-human animal wherein the seal formulation is nontoxic heavy metal salt such as bismuth (U.S. Pat. No. 8,353,877); a method of applying to the teat canal and/or teat sinus a composition comprising exogenous keratin (U.S. Pat. No. 8,226,969); a method of forming a physical barrier in the teat canal for prophylaxis during an animal's dry period by infusing an amount of a teat seal formulation into the teat canal of the animal, wherein the teat seal formulation comprises a bismuth-free, nontoxic, heavy metal salt of titanium, zinc, barium or combinations thereof and the physical barrier does not cause a black spot defect in dairy products made with milk from the animal (U.S. Pat. No. 7,906,138); a method of forming an anti-infective free physical barrier in the animal's teat canal for prophylactic treatment of mastitis during the dry period comprising the step of infusing a seal formulation into the teat canal of the animal without an anti-infective agent, wherein the seal formulation comprises a nontoxic heavy metal salt such as bismuth in a gel base of aluminum stearate with a vehicle such as liquid paraffin or a gel base comprising a polyethylene gel (U.S. Pat. No. 6,254,881) and the like.
However, none of the existing seal formulations or external dip products seals the teat of the dairy animal externally for a sufficient amount of time to prevent mastitis, particularly the form that can be fatal and/or very contagious in the animals, like among heifers. Moreover, while teat sealants have been established as a viable method to provide a higher level of protection regardless of antibiotic choice or administration, the current products on the market fail to meet the demand for ease of use and long-lasting tissue adherence, ease of removal, avoidance of milk contamination and prevention of black spot defect in aged cheese. What is needed, therefore, is a nontoxic formulation that is easy and safe for the animal handler to administer and that preferably forms an effective, long-lasting seal in place directly on the tissue (that is, “in situ”). Additionally, it is necessary for the seal formulation not to interfere with the quality of the dairy animal's milk, yogurt or cheese products created from the milk, especially for the sealant to avoid the black spot defect in aged cheese. Indeed, there is a definite art-recognized need in the veterinary field to find a long-lasting, nontoxic, non-irritating seal formulation that forms an adequate barrier on the animal's teat to prevent or to reduce significantly the incidence of mastitis caused by pathogens, preferably without the use of antibiotics or other medicinal agents that require a withholding period for public consumption of the animal's milk. There is also a definite need to find a long-lasting seal formulation that can contain antibiotics and the like for the effective treatment or prevention of mastitis.
It is an object of the present technology, therefore, to provide sol-gel polymer composites that ameliorate the inconveniences of the known hydrogels.